The present invention relates to methods and devices useful for analytical testing. Such testing includes, but is not limited to medical diagnosis and environmental testing.
The following is a discussion of relevant art, none of which is admitted to be prior art to the present invention.
A flow-through, or porous, assay device is described in U.S. Pat. No. 4,632,901 by Valkirs, et al. In this method an immunoassay is performed on a membrane or filter which is coated with an antibody and is capable of removing an analyte from a sample applied to the membrane. Visualization is based on the analyte dependent capture of a secondary reagent which will act on a substrate and produce a colored, particulate product which will non-specifically adhere to the membrane only where the secondary reagent is present. Numerous modifications to this basic design have been introduced including colored, and/or metallic particles (U.S. Pat. No. 4,775,636) attached to the secondary reagent for visualization, and the introduction of chromatographic rather than flow-through techniques (U.S. Pat. No. 5,232,835).
U.S. Pat. No. 5,200,312 describes a membrane assay system where a colored, insoluble product is used for the detection of an analyte. This product is formed by an enzyme interacting with a substrate that contains a reagent which when exposed to the enzyme produces a chromophore containing insoluble product producing a visible color change. U.S. Pat. No. 5,395,754 describes methods for producing control or calibration zones on a membrane surface for use in a biological assay.
Production of porous antireflective films have been described (66 J. Opt. Soc. Am. 515-519, 1976; 66 J. Am. Ceramic Soc. 302-307, 1983). The antireflective films have steep refractive index gradients for making broad band AR layers. The films are highly porous with the pores being disordered and interconnected. The pores capture air within the AR material being formed which help produce a refractive index gradient.
Mass transport, or mass transfer, is a well established phenomena. It can arise from the presence of a concentration gradient, temperature gradient, electrical field, gravity, etc. Mass transport in a solution is very sensitive to solution movement or flow or convection. Mass transport may also be influenced by the diffusion coefficient or charge of materials in the solution.
In a static diffusion limited reaction, a concentration gradient can be formed as the diffusion layer is depleted and the analyte concentration is reduced at the surface. Analyte from a higher concentration zone in the sample must diffuse to the surface for binding. Only sample near the surface will be bound. Replenishing analyte to the diffusion layer or barrier limits the binding reactions. Convective mass transport effects can serve to disrupt or modify the diffusion barrier.
Solution flow, mass transport, in a highly porous or interconnected surface is turbulent, producing plug or convection flow characteristic. However, in a channeled surface, the hydrodynamic mass transport creates laminar flow characteristics. Plug flow causes the solution to mix by convection and then advance along its path. This ensures that the diffusion barrier is minimized as sample flows laterally across the porous material. In an assay system, plug flow could increase the probability of non-specific adhesion of non-analyte material and subsequent visualization reagents. However, the convective flow will tend to increase the contact of analyte with available binding sites as the flow path is followed by fresh solution which repeatedly contacts the available binding sites.
Solutions which flow through or across channeled material are essentially static when in contact with a solid, uniform surface until a channel is encountered. Flow through that channel creates laminar flow. Thus, while a reaction is diffusion limited, material flow is influenced such that the diffusion barrier or layer is disrupted. The convection introduced by channels continuously forces new analyte to the surface eliminating the dead layer near the pore. While, also preventing the formation of a diffusion barrier which meets the static condition between the pores. Thus, the laminar flow continuously brings new bulk into the diffusion boundary. It is commonly believed that the plug flow system is more efficient in overcoming the diffusion limitation than the laminar flow system. Applicant has suprisingly discovered that for the optical assay devices of the present invention laminar flow is more effective than plug flow systems.
In a static solution/solid reaction, the diffusion barrier, after 20 seconds, is δ (t)=2.8×10−3 cm (δ(t)=2 (Dot)−1/2). Do is assumed to be 1×10−7 cm2/sec for common biologicals. In a hydrodynamic mass transport case, the diffusion barrier is essentially independent of time and δ (o)=3.7×10−4 cm (δ(o)=1.61 (Do)1/3 (ωv1/6)−1/2) Where ω is the angular frequency based on a solution moving across an assumed solid having an angular velocity of ω and v is a function of the solutions viscosity (kinematic viscosity). The value of v based on a solution moving across an assumed solid having an angular velocity of ω was assumed to be 0.01 cm2 sec−1 (water). Calculations are derived from Ficks Law.